Fig. 7
From: ERAP1 promotes Hedgehog-dependent tumorigenesis by controlling USP47-mediated degradation of βTrCP
a–e SHH-MB PDX tumor cells were treated with different amounts of Leu-SH. a, b Cells were counted with trypan blue at the indicated time points to evaluate the growth rate of viable cells (a) and the percentage of cell death (b). c Cell survival was also assessed by the Cell Titer Glo assay. d Immunofluorescent staining of cleaved Caspase 3 (green) of Leu-SH treated cells (48 h). Nuclei were counter stained with Hoechst (Blue). Scale bar 5 μm. e Quantification of cleaved Caspase-3 from immunofluorescence stainings in (d). (f–k) NSG mice (n = 6 for group) were grafted with SHH-MB PDX tumor cells. Tumor masses (150 mm3) were intratumorally injected with Leu-SH (0.528 μM/Kg). f Tumor growth was monitored every three days. g Quantification of the explanted tumor masses (upper panel) and representative flank xenograft average volumes (lower panel). h H&E, Ki67, NeuN and cleaved Caspase-3 immunohistochemical staining of PDX tumor samples. i Quantification of Ki67, NeuN and cleaved Caspase-3 from immunohistochemical stainings in h. Scale bar 100 μm. j, k mRNA (j) and protein (k) expression levels of the indicated Hh target genes from the tumor masses assayed in g. Mean ± S.D. *P < 0,05; **P < 0,01; ***P < 0,001; ****P < 0,0001 determined using two-tailed Student’s t-test
